Biomedical subjects

T C Wang

Publications and source records attributed to T C Wang.

At least 109 records · Page 6Linked to original sources

PubMed1996

Rat histidine decarboxylase promoter is regulated by gastrin through a protein kinase C pathway.

The enzyme L-histidine decarboxylase (HDC; EC 4.1.1.22), which converts L-histidine to histamine, plays a key role in the regulation of acid secretion. In the rat and human stomach, the peptide hormone gastrin appears to be one of the main regulators of HDC expression. In rats, marked elevation of gastric HDC mRNA abundance was observed within 12 h after induction of hypergastrinemia by a single injection of the proton-pump blocker omeprazole. In situ hybridization revealed that HDC expression occurred in the basal third of gastric glands where enterochromaffin-like cells are localized. To study the regulation of HDC gene transcription, 1,291 nucleotides of the 5'-flanking region of the rat HDC gene and the noncoding portion of exon 1 were cloned and sequenced. Gastrin and cholecystokinin (CCK) octapeptide equipotently stimulated the transcriptional activity of the rat HDC promoter three- to fourfold, and deletion analysis revealed the presence of a gastrin response element within 201 nucleotides upstream of the translational start site. Time-course studies revealed maximal activation of the HDC promoter after 12-36 h. Direct stimulation of protein kinase C (PKC) with the phorbol ester phorbol 12-myristate 13-acetate (PMA) substantially elevated rat HDC promoter activity, whereas induction of Ca2+ -dependent signaling pathways with thapsigargin was without effect. Downregulation or blockade of PKC abolished the effects of gastrin and PMA on the HDC promoter. These data indicate that stimulation of the CCK-B/gastrin receptor activates the rat HDC promoter in a time- and dose-dependent fashion and that this effect is primarily mediated via a PKC-dependent signaling pathway. Use of HDC as a model gene will allow further investigation of the intracellular pathways that are involved in gastrin-dependent gene regulation.

Animals↗

PubMed1996

2-D time-of-flight (TOF) MRA of thrombophlebitis of upper extremity and subclavian veins. A case report.

In this report, the authors present a case of segmental thrombosis of left subclavian vein that was inaccessible to conventional venography and inconclusive in magnetic resonance techniques (spin echo, gradient echo) but was clearly demonstrated by two-dimensional time-of-flight magnetic resonance angiography (2D TOF MRA). This technique is very sensitive to detect slow flow or partial occlusion of the vessels and is of great help in solving the problem of axillary-subclavian venous obstruction or thrombophlebitis.

Adult↗

PubMed1996

Synthesis and evaluation of coumarin alpha-methylene-gamma-butyrolactones: a new class of platelet aggregation inhibitors.

In a search for new inhibitors of platelet aggregation, certain coumarin-bearing alpha-methylene-gamma-butyrolactones were synthesized and evaluated for inhibitory activity against thrombin (Thr)-, arachidonic acid (AA)-, collagen (Col)-, and platelet-activating factor (PAF)-induced aggregation in washed rabbit platelets. These compounds were efficiently synthesized from commercially available 7-hydroxycoumarin or its derivatives. Among them, 7-[(2,3,4,5-tetrahydro-4-methylene-5-oxo-2-phenyl-2-furanyl) methoxy]-2H-1-benzopyran-2-one (3d) showed the most potent inhibition of AA- and PAF- induced aggregation, with IC50 values of 3.65 and 16.36 microM respectively.

Adenosine Triphosphate↗

PubMed1995

Molecular cloning and sequencing of the murine gastrin gene.

Gastrin is a peptide hormone important in acid regulation, growth of enterochromaffin cells of the oxyntic mucosa, and smooth muscle contractility. We isolated a genomic clone of gastrin from a SV-129 murine genomic library. The murine gastrin gene contains an open reading frame of 101 amino acids. The deduced amino acid sequence has 91.1% homology with the rat and 67.3% homology with the human analogue. Southern blot analysis of the murine gastrin clone gave the same restriction pattern as that seen from mouse kidney genomic DNA, consistent with this being a single copy of the gastrin gene in the mouse genome. Transfection studies demonstrated that the murine gastrin gene expression is stimulated by epidermal growth factor to the same extent as the human gastrin gene.

Amino Acid Sequence↗

PubMed1995

Purification and identification of pituitary cytotropic factor.

It has been shown that the pituitary contains a cytotropic factor (CTF) that stimulates the secretion of catecholamines by dopaminergic neurons of the hypothalamus. In the present study, CTF was purified from rat pituitaries and found by means of mass spectrometric analysis to be adenosine. This finding was corroborated by the observations that CTF behaves identically to adenosine when subjected to liquid chromatography, is inactivated and converted to inosine by adenosine deaminase, and is qualitatively and quantitatively indistinguishable from adenosine in its biological activity. It is concluded that pituitary adenosine is a trophic factor for hypothalamic dopaminergic neurons.

Adenosine↗

PubMed1995

Structure of a retro-binding peptide inhibitor complexed with human alpha-thrombin.

The crystallographic structure of the ternary complex between human alpha-thrombin, hirugen and the peptidyl inhibitor Phe-alloThr-Phe-O-CH3, which is acylated at its N terminus with 4-guanidino butanoic acid (BMS-183507), has been determined at 2.6 A resolution. The structure reveals a unique "retro-binding" mode for this tripeptide active site inhibitor. The inhibitor binds with its alkyl-guanidine moiety in the primary specificity pocket and its two phenyl rings occupying the hydrophobic proximal and distal pockets of the thrombin active site. In this arrangement the backbone of the tripeptide forms a parallel beta-strand to the thrombin main-chain at the binding site. This is opposite to the orientation of the natural substrate, fibrinogen, and all the small active site-directed thrombin inhibitors whose bound structures have been previously reported. BMS-183507 is the first synthetic inhibitor proved to bind in a retro-binding fashion to thrombin, in a fashion similar to that of the N-terminal residues of the natural inhibitor hirudin. Furthermore, this new potent thrombin inhibitor (Ki = 17.2 nM) is selective for thrombin over other serine proteases tested and may be a template to be considered in designing hirudin-based thrombin inhibitors with interactions at the specificity pocket.

Amino Acid Sequence↗

PubMed1995

DNA strand breakage, thymine glycol production, and hydroxyl radical generation induced by different samples of crystalline silica in vitro.

Five preparations of alpha-quartz [Min-U-Sil 5 (MQZ), MQZ pretreated with hydrofluoric acid (HFMQZ), Chinese standard alpha-quartz (CSQZ), and two German samples, DQ-12 and F600] and two preparations of the crystalline silica polymorphs, cristobalite and tridymite, previously characterized for surface area and surface charge, were evaluated for their relative activities in the following assays: (i) in vitro assays of short duration (< or = 15 min) for oxygen consumption and for generation of hydroxyl radicals (measured by electron spin resonance spin trapping), and (ii) in vitro assays of longer duration for DNA strand breakage (measured using linear DNA as a detector molecule) and for production of the oxidized DNA base, thymine glycol (measured by gas chromatography-mass spectrometry). Marked differences among the samples were found for their levels of oxygen consumption and of hydroxyl radicals' generation. All samples caused increased formation of thymine glycol, with wide variations in activity among samples. When normalized for equal surface area, the samples produced different levels of DNA strand breakage. Addition of hydrogen peroxide strongly accelerated DNA damage--more for cristobalite than for the alpha-quartz samples. DNA damage by quartz was enhanced by ferric chloride and inhibited by iron chelators. The order of relative activity of the samples varied with different types of in vitro assays and was not directly correlated to surface area. Electrophoretic mobility, as measured by zeta potential, was not significantly different among samples. The results suggest that the ability of different crystalline silica samples to generate a rapid burst of oxygen free radicals is distinct from their ability to induce DNA damage and DNA base oxidation over longer time periods. The relative activities of the samples in cellular assays (hemolysis of human erythrocytes; cytotoxicity and neoplastic transformation of BALB/3T3/A31-1-1 cells) were in turn markedly different from those listed above, suggesting a more critical role for surface area. The mechanisms of carcinogenesis by crystalline silica need to be further investigated in relation to the underlying physicochemical characteristics.

Animals↗

PubMed1995

A rat gastrin-human gastrin chimeric transgene directs antral G cell-specific expression in transgenic mice.

Gastrin gene expression in the gastrointestinal tract is under both developmental and spatial regulation. In the mature animal, gastrin, an important regulator of parietal acid secretion, is expressed primarily in G cells of the antrum. To determine whether specific promoter elements can direct expression to the gastric antrum in vivo, 450 nucleotides of the proximal rat gastrin promoter were cloned and used to construct a rat gastrin-human gastrin reporter chimeric transgene, which was injected into the mouse germ line. Northern blot analysis, in situ hybridization, and double-label immunocytochemistry studies demonstrated expression of the transgene specifically in antral G cells. Low levels of transgene expression were observed in the ileum and colon, where immunohistochemical studies demonstrated colocalization in enteroendocrine cells expressing peptide YY. The same 450-nucleotide rat gastrin promoter, when joined to the human growth hormone gene, did not result in antral expression. Similarly, a human gastrin-human gastrin reporter transgene also did not achieve antral expression, although it did express in the liver. These results suggest that cis-acting elements present in both the basal 450-nucleotide rat gastrin promoter and the intragenic sequences of the human gastrin gene are necessary to direct expression of a transgene specifically to antral G cells.

Amino Acid Sequence↗

PubMed1995

Transforming growth factor alpha disrupts the normal program of cellular differentiation in the gastric mucosa of transgenic mice.

Transforming growth factor alpha (TGF alpha) evokes diverse responses in transgenic mouse tissues in which it is over-expressed, including the gastric mucosa, which experiences aberrant growth and a coincident repression of hydrochloric acid production. Here we show that ectopically expressed TGF alpha induces an age-dependent cellular reorganization of the transgenic stomach, in which the surface mucous cell population in the gastric pit is greatly expanded at the expense of cells in the glandular base. Immunohistochemical analysis of BrdU incorporation into DNA demonstrated that although mature surface mucous cells were not proliferating, DNA synthesis was enhanced by approximately 67% in the glandular base and isthmus, where progenitor cells reside. RNA blot and in situ hybridization were employed to determine temporal and spatial expression patterns of specific markers representing a variety of exocrine and endocrine gastric cell types. Mature parietal and chief cells were specifically depleted from the glandular mucosa, as judged by a 6- to 7-fold decrease in the expression of genes encoding H+,K(+)-ATPase, which is required for acid secretion, and pepsinogen C, respectively. The reduction of these markers coincided in time with the activation of TGF alpha transgene expression in the neonatal stomach. The rate of cell death in the glandular region was not overtly different. Significantly, the loss of parietal and chief cells occurred without a concomitant loss of their respective cellular precursors. In contrast to exocrine cells, D and G endocrine cells were much less severely affected, based on analysis of somatostatin and gastrin expression. Analysis of these dynamic changes indicates that TGF alpha can induce selective alterations in terminal differentiation and proliferation in the gastric mucosa, and suggests that TGF alpha plays an important physiological role in the normal regulation of epithelial cell renewal.

Animals↗

PubMed1995

Evaluation of iliopsoas compartment disorders by computed tomography.

BACKGROUND: Iliopsoas compartment lesion frequently goes undiagnosed at clinical presentation. Therefore, even delayed diagnosis may depend on radiographic techniques. Computed tomography (CT) can play an important role in the identification and localization of these lesions. This report concerns the experience of using CT to evaluate the iliopsoas compartment disorder. METHODS: Retrospective review of CT scans for 42 patients who had abnormalities of the iliopsoas compartment revealed that inflammation was responsible in 20, hemorrhage in 8, and tumor in 14 patients. Diagnoses were proven by surgery or biopsy/aspiration in all cases except in five cases whose CT findings correlated with their clinical pictures. A comparison was made among the surgical data, the CT and plain abdominal films. In addition, a comparative evaluation of CT features was also made. RESULTS: By using a hypodense area for the diagnosis of psoas abscess, the sensitivity, specificity and accuracy of CT were 100%, 77%, and 88%, respectively. By using a diffuse involvement of the entire muscle for the diagnosis of hemorrhage, the sensitivity, specificity and accuracy of CT were 63%, 62%, and 62% respectively. By using a bone destruction for the diagnosis of tumors, the sensitivity, specificity and accuracy of CT were 79%, 39%, and 69%, respectively. Evaluation of the iliopsoas compartment lesions was 91% sensitive by CT and 40% sensitive by plain X-ray film. CONCLUSIONS: It is difficult to differentiate iliopsoas abscesses, hematomas and neoplasms without knowing the clinical history. However, when coupled with percutaneous aspiration biopsy, CT has proved to be an effective, rapid means to diagnose and guide the therapy of psoas lesions.

Adult↗

PubMed1994

Liquid chromatography/mass spectrometry of phospholipids using electrospray ionization.

An improved technique for phospholipid molecular species analysis was developed using high-performance liquid chromatography/mass spectrometry with the electrospray interface. Using the 0.5% ammonium hydroxide in a water-methanol-hexane mixture and a C-18 column, complex mixtures of phospholipid molecular species were separated and detected mainly as protonated or natriated molecular species. The response was linear over 2 orders of magnitude, allowing quantification of each molecular species. In comparison to the existing LC/MS techniques, marked improvement in sensitivity was observed. The present quantification limit is approximately 0.5 pmol before split (5 fmol after 1/100 split). The relative responses were more dependent on the head group identity rather than fatty acyl composition within a phospholipid class. In general, phosphatidylcholine (PC) species are most sensitively detected followed by phosphatidylethanolamine (PE) species. The sensitivity of phosphatidylserine (PS) in the positive ion mode is approximately 20 times less in comparison to PC under our condition.

Animals↗

PubMed1994

Differential proliferative responses of Syrian hamster embryo fibroblasts to paraquat-generated superoxide radicals depending on tumor suppressor gene function.

Oxygen radicals have been widely implicated in neoplastic transformation; however, little is known regarding their mode of action. In an attempt to delineate potential mechanisms of action, an analysis of superoxide effects on cell growth was studied in normal and two nontumorigenic, immortal cell lines derived from normal Syrian hamster embryo (SHE) fibroblasts. The two immortal cell lines differed in their ability to suppress tumorigenicity of tumor cells in cell hybrids. One cell line suppressed tumorigenicity (sup+), while a second clone was unable to suppress tumorigenicity (sup-). Paraquat was used to generate superoxide through its capacity to be reduced by NAD(P)H and to generate superoxide radicals. The growth response of the various cell types was measured by colony-forming ability as well as by tritiated thymidine incorporation using autoradiography. At low paraquat concentrations (25 microM), primary SHE cells and two sup+ clones showed up to a 40% enhancement in colony formation, while two sup- clones showed no increase. Toxicity was observed at high doses, starting at approximately 100 microM paraquat. Since oxygen radicals are also mutagenic, primary SHE cells were examined for chromosomal aberrations. Chromatid gaps and breaks were induced at all concentrations of paraquat used. Thus, superoxide not only causes cellular toxicity at high doses but at low doses enhances cell growth of certain cells (primary SHE cells and sup+ cells) but not others (sup- cells). Therefore, differing responses of cells at different stages of neoplastic progression must be considered in understanding oxygen radical effects in growth control and carcinogenesis.

Animals↗

PubMed1994

Mammary hyperplasia and carcinoma in MMTV-cyclin D1 transgenic mice.

Physical associations between cyclins, viral oncogenes and tumour suppressor genes imply a central role for cyclins in growth control. Cyclin D1 was identified as a candidate oncogene (PRAD1) in tumour-specific DNA rearrangements and is suspected to be a contributor to several types of neoplasms including breast cancer. Cyclin D1 also rescues G1 cyclin-defective Saccharomyces cerevisiae, and is a growth-regulated gene. Despite evidence suggesting that cyclin D1 is an oncogene, its ability to transform cells directly in culture remains controversial. To evaluate its potential to deregulate growth in vivo in a physiologically relevant tissue we overexpressed cyclin D1 in mammary cells in transgenic mice. We report here that overexpression of cyclin D1 resulted in abnormal mammary cell proliferation including the development of mammary adenocarcinomas. We conclude that overexpression of cyclin D1 deregulates cell proliferation and can induce tumorigenic changes in mammary tissues, suggesting that cyclin D1 indeed plays an important oncogenic role in breast cancer.

Adenocarcinoma↗

PubMed1994

Okazaki DNA fragments contain equal amounts of lagging-strand and leading-strand sequences.

The question of whether discontinuous DNA replication operates only for the lagging strand or for both strands in Escherichia coli is still not resolved. Most in vitro data indicate that DNA replication is semi-discontinuous. In contrast, the in vivo data indicate that DNA replication is discontinuous in both strands. Proponents for the semi-discontinuous DNA replication model argued that the failure of demonstrating semi-discontinuous DNA replication in vivo may be due to the breakdown of a continuously synthesized strand by DNA repair processes. In this article, we present further evidence that the apparent discontinuous DNA replication in E. coli is not the result of DNA repair processes. Furthermore, we show that the Okazaki fragments contain about equal amounts of leading- and lagging-strand sequences. These results are most consistent with the discontinuous DNA replication model as originally proposed by Okazaki et al.

Autoradiography↗

PubMed1994

Activation of gastrin transcription in pancreatic insulinoma cells by a CACC promoter element and a 70-kDa sequence-specific DNA-binding protein.

Gastrin gene expression in the pancreatic islets is developmentally regulated and occurs largely during fetal life. Deletional analysis of transiently transfected rat insulinoma cells with gastrin 5'-flanking sequences in luciferase reporter genes demonstrated that the gastrin promoter sequence proximal to -111 base pairs (bp) contains the cis-regulatory elements necessary for maximal transcription. Mutational analysis identified the sequence CCCCACCCCA (-109 to -100 bp) as a positive cis-regulatory element (CACC) located 5' to a previously described negative element (-100 to -90 bp) and E-box positive element at -82 bp. Multimers of the CACC element in a heterologous promoter activated transcription independent of the other cis-regulatory elements. CACC binding proteins were purified from insulinoma cell nuclear extracts by cation exchange and affinity chromatography. Southwestern blot of nuclear extracts identified a 70-kDa CACC-binding protein. Mutational analysis of the CACC element showed a close correlation between DNA binding of this protein and transcriptional activation. Transcriptional activation by multimers of the CACC element in a heterologous promoter was detected in a variety of cell lines but was strongest in those of islet lineage. Likewise, the presence of the 70-kDa CACC-binding protein was found in many cell lines but was most abundant in the insulinoma cells. The CACC-binding protein has not been previously identified among the known pancreatic regulatory factors and may have an important role in the developmental expression of gastrin.

3T3 Cells↗

PubMed1994

Purification and kinetic mechanism of the glutathione S-transferase from C6/36, an Aedes albopictus cell line.

Glutathione S-transferase from an Aedes albopictus cell line, C6/36, was purified to apparent homogeneity by a single glutathione-Sepharose 4B affinity chromatography, with an overall yield of 66% and 226-fold purification. The enzyme is presumably a homodimer with subunit M(r) 23,000, which is similar to the enzyme isolated from other sources. Under native conditions, the enzyme exhibited multiple forms upon isoelectric focusing or polyacrylamide gel electrophoresis, and all these forms retained enzymatic activity. The pI value of the purified enzyme was distributed between approximately 4.7 and 5.4 with major form at 4.9. The purified enzyme lost 63% activity at -20 degrees C within 1 week. Stability of the purified enzyme was greatly improved by the addition 10 mg/ml of bovine serum albumin, under which only 7% activity was lost after 1 week at -20 degrees C. Activation energy for the enzyme-catalyzed conjugation of glutathione with 1-chloro-2,4-dinitrobenzene was found to be 49.1 kJ/mol. The enzyme could utilize 1-chloro-2,4-dinitrobenzene and ethacrynic acid as substrate, but not bromosulfophthalein, cumene hydroperoxide, or trans-4-phenyl-3-buten-2-one. The initial-velocity and product-inhibition studies indicated that the enzyme reaction conformed to a steady-state random Bi-Bi kinetic mechanism, which was similar to the glutathione S-transferase from other sources. The kinetic data also indicated a synergistic effect between the binding of glutathione G-site and that of organic electrophilic substrate in the H-site of the enzyme active center. The biological significance of the conjugation reaction is discussed.

Aedes↗

PubMed1994

Spasmolytic polypeptide: a trefoil peptide secreted by rat gastric mucous cells.

BACKGROUND/AIMS: Spasmolytic polypeptide (SP) is a trefoil peptide expressed in the digestive tract. This study aimed to determine the structure and distribution of SP expression in the rat gastrointestinal tract. METHODS: The structure of rat SP was determined from the sequence of complementary DNAs isolated from antral RNA. SP gene expression was localized by Northern blotting and in situ hybridization in the adult and fetal rat digestive tract. Expression of the SP peptide was localized by immunocytochemistry and Western blot analysis. RESULTS: SP messenger (m)RNA was found predominantly in the stomach with highest expression in the antrum. High levels of SP mRNA were expressed in the fetal stomach before gastrin and somatostatin expression. Surprisingly, SP mRNA and peptide did not colocalize in the gastric mucosa, SP mRNA being superficial to SP peptide immunoreactivity throughout the gastric mucosa. Abundant SP immunoreactivity was seen in the lumen of the gastric glands and the mucus layer adherent to the gastric mucosa, indicating luminal secretion. CONCLUSIONS: In the rat, SP is a peptide secreted predominantly from antral mucous cell. The high concentrations of SP in the adherent gastric mucus layer (approximately 10 mumol/L) suggest that SP functions as a structural peptide rather than a regulatory peptide.

Amino Acid Sequence↗

PubMed1994

Involvement of RecF pathway recombination genes in postreplication repair in UV-irradiated Escherichia coli cells.

Mutations affecting the RecF pathway of recombination (recF, recG, recJ, recN, recO, recQ, recR, ruvA, ruvC) were systematically introduced into two sets of strains: (a) uvrA and uvrA recA2020, (b) uvrA recBC sbcBC and uvrA recBC sbcBC recA2020. We examined: (i) the effect of these mutations on the repair of DNA daughter-strand gaps which are produced in the nascent DNA synthesized after UV irradiation, and (ii) the ability of recA2020 (a suppressor for the recF mutation) to suppress the UV radiation sensitivity caused by these mutations. In the uvrA cells, mutations in recF, recR or recO genes produced a major deficiency in the repair of daughter-strand gaps, whereas mutations in recJ, recG, recN, recQ, ruvA or ruvC genes had no effect on the repair of daughter-strand gaps. In both uvrA and uvrA recBC sbcBC backgrounds, the UV radiation sensitivity caused by recF, recG, recR, recO, ruvA, or ruvC mutations was partially suppressed by recA2020, whereas the UV radiation sensitivity caused by recJ, recN, or recQ mutations was not suppressed by recA2020. Partial suppression of the UV sensitivity of recG, ruvA and ruvC mutants was not observed with other suppressors for recF, i.e., recA441, recA720 and recA730. Taken together, these results further support the notion that the recF, recR and recO gene products (abbreviated as RecFOR) function at the same step in recombination repair, possible as a complex. It also suggests that this putative RecFOR complex does not contain proteins encoded by other genes involved in the RecF pathway of recombination.

Alleles↗